1. Field of the Invention
The present invention relates generally to methods and apparatus for determining analytes in biological specimens. More particularly, it relates to the construction and use of a liquid sample applicator having an internal matrix which removes particulates from the sample and to a liquid sample applicator having a permeable internal filter with a reagent dispersed therein.
2. Description of the Background Art
The immunoassay of biological specimens has proved to be of enormous benefit in a variety of circumstances, such as the diagnosis of disease, detection of drugs, and monitoring of physiological metabolites. In performing immunoassays on liquid biological samples, pretreatment is frequently required to remove particulates, such as cellular debris, which might foul the assay apparatus or interfere with the assay results. The particulates may be removed, for example, by filtering the liquid sample during a separate step in the assay procedure. Although effective to overcome the interference problems, the addition of a filtering step is undesirable as it adds to the time and complexity of the assay procedure. It would therefore be desirable to provide filtering procedures and apparatus which would be integral to the assay protocol and would not add time or complexity to the overall procedure.
One attempt to simplify the filtering of biological samples undergoing immunoassay procedures is described in U.S. Pat. No. 3,873,682, to Ogawa. A filter cartridge is detachably mounted on the tip of a pipette or fluid dropper. By drawing sample upward through the filter into the pipette, the particulates are removed. The filter is discarded prior to discharging the sample into an appropriate assay media. Although effective in removing particulates, the device of Ogawa is relatively expensive as it consists of two separate pieces. Moreover, the method requires an extra step to discard the cartridge. In addition to extending the time required to perform the assay, the need to discard the filter cartridge would complicate use of the pipette in most automated systems for performing immunoassays. Pipettes manufactured in accordance with the patent are commercially available from Mochida Seiyaka Kabushiki Kaisha, Tokyo, Japan.
Pipettes or dispensers containing an integral filter are disclosed in U.S. Pat. Nos. 4,483,825 (to Fatches) and 4,487,696 (to Ferrara). These patents are directed to the separation of blood cells from plasma in centrifuged blood samples. Each discloses a tube having an inlet end containing a filter through which the plasma is passed. However, the filtered serum is then poured out of the tube through the opposite, or outlet end. While such a method would remove particulates, it would be necessary to remove the bulb from the top of the fluid dropper prior to dispersing the filtered liquid, thus adding an extra step and additional time to the assay process. Additionally, dispersing the fluid from the opposite end of the tube would be unworkable in most if not all automated immunoassay systems and with one-piece pipettes.
A different approach for sample filtration is found in the TestPack.RTM. assay system available from Abbott Laboratories, North Chicago, Ill., and the Preview Serum/Urine-hCG assay system available from Leeco Diagnostics, Inc., Southfield, Michigan. In both cases, a detachable filter is snapped into place against a membrane having bound antibodies specific for the analyte of interest. Sample is applied to the membrane through the filter, and the filter removed prior to development of the assay. Although functional in removing particulates, use of such assay systems has certain disadvantages. In particular, the time required for the sample to penetrate the filter can be as long as one minute or more, adding to the time necessary to perform the assay. Moreover, the need to remove the filter holder prior to development is an extra step which further extends the time required to perform the assay and which, if forgotten, can ruin the test results completely.
For these reasons, it would be desirable to provide a method and apparatus for filtering biological specimens during immunoassay procedures, which methods and apparatus do not require any additional steps or procedures, which do not extend the time required to perform the assay, and which still provide the improved assay results associated with the more cumbersome filtering techniques described above.
Additionally, biological assays often require the combination of one or more reagents with a test sample or other liquid assay component during the course of the assay. Heretofore, reagent addition has generally required discrete addition step(s) performed at appropriate times during the assay. While certainly workable, each additional assay step increases the assay complexity, the time required to perform the assays, and the chance that contaminants and errors will be introduced into the final result. It would therefore be desirable to perform reagent combination(s) concurrently with the transfer of liquid sample or other liquid assay component, thus reducing the number of actual assay steps involved, with a concommitant reduction in time and effort required to perform the assay.